Transfection of Theileria parva and the role of genes encoding QP-rich proteins in host-parasite interactions

This proposal aims at developing stable transfection in T. parva through a systematic approach
starting with transient transfection, identification of suitable promoter
regions and the development of suitable selectable markers to be used in
integration plasmids. The use of a new generation transfection device and the
expertise in large sporozoite stabilate production should contribute
significantly to its success.

The transfection of T. parva
sporozoites will enable us to investigate several aspects of this unique
host-parasite interaction. The project will focus on the function of genes
encoding QP-rich proteins.  Some insight
gained from previous research and a set of preliminary results related to these
genes is already available, but several important questions remain that can
only be addressed through transfection experiments. In particular the pattern
of expression and the role of PIM and the SVSP gene family in host-parasite
interactions will be further explored. This will involve the construction of
plasmids allowing the stable expression of fluorescently tagged parasite
proteins. This will enable a detailed exploration of protein localization and
interactions with host cell structures, providing important clues on the
potential function of these genes in Theileria-transformed
cells.

In a later phase, various partial gene deletion constructs will be
generated to investigate in detail the expression and interaction of specific
parasite protein domains with host cell components or cellular structures. The
possibility of using stably expressed tagged forms of PIM, SVSP or other
candidate proteins in the search for host cell proteins that interact with
these parasite proteins in their ‘natural’ context will also be explored.

The use of fluorescently tagged parasites opens up the possibility to
look at parasite differentiation and monitor the influence of drugs like tetracycline
on parasite development in in vitro
cultures. In the long run, this research might open up novel approaches towards
immunization against this important tropical cattle pathogen.